Atrial fibrillation (AF) is the most common cardiac arrhythmia. Its prevalence doubles with every decade of life, exceeding 5% in people older than 65. AF is associated with a 5-6 fold increase in the incidence of stroke, due almost exclusively to thrombus formation in the left atrium. Stroke risk is increased by over 17 fold in patients with rheumatic mitral valve disease-related AF. In the elderly, AF is the major cause of thromboembolic events; which in turn drastically increases mortality. It has become clear recently that AF patients possess a hypercoaguable state but with unclear molecular mechanisms. One (1) possible explanation is that the anti-coagulating defense mechanisms were deficient in AF. Tremendous amount of work has well documented the anti-inflammatory and anti-thrombotic properties of nitric oxide (NO) in the vascular endothelium. Likewise, NO protects the integrity of endocardial function. Thus loss of endocardial NO may represent a novel molecular mechanism whereby hypercoaguable state develops in patients with AF. Indeed, our recent study for the first time demonstrated that NO bioavailability was reduced in the endocardium in a porcine model of AF, whereas the pro-thrombotic protein plasminogen activator inhibitor 1 was upregulated. To further characterize this response in humans, we propose to determine whether endocardial dysfunction, characterized by a loss of NO and mis-regulation of proteins modulating thrombotic potential, occurs in patients with AF (aim 1). In aim 2, we will determine effects of AF on endocardial NO synthase (eNOS) signaling and cardiac cGMP production. Recent studies have shown that human AF is associated with increased production of reactive oxygen species (ROS), which in turn rapidly inactivate NO. Furthermore, oxidation of eNOS cofactor tetrahydrobiopterin causes dysfunction of eNOS where the enzyme produces ROS rather than NO. This phenomenon has now been referred to eNOS uncoupling and could potentially prolong endocardial dysfunction in AF. Therefore, in aim 3 we will determine whether uncoupled eNOS contributes to ROS production in AF and whether tetrahydrobiopterin administration improves endocardial function in patients with AF. Overall this project is designed to investigate whether and how endocardial dysfunction occurs in humans with AF. Because endocardial dysfunction likely predicts AF patients to thromboembolic events, these studies would eventually lead to novel therapeutic modalities targeting endocardial dysfunction. These modalities are anticipated to reduce stroke risk in patients with AF which in turn would markedly reduce modality in the elderly.